Methods of predicting susceptibility to infectious disease and related methods of treatment

ABSTRACT

Disclosed herein are methods for predicting susceptibility to an infection based on the observation of specific mutations in blood cells, as well as methods for treating or reducing susceptibility to an infection in a subject.

RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.17/240,991, filed on Apr. 26, 2021, which claims the benefit of U.S.Provisional Application No. 63/015,104, filed on Apr. 24, 2020. Theentire teachings of the above applications are incorporated herein byreference.

BACKGROUND OF THE INVENTION

Coronavirus disease 2019 (COVID-19) became a pandemic in early 2020. Oneof the reasons for its rapid and widespread diffusion is the presence ofa large number of asymptomatic or paucisymptomatic patients, who are notrecognized as infected and therefore contribute to the spread of thevirus. Data on the spread of the virus have led scientists to think thatthe actual number of infected people is much higher than the number ofpositive tests. For example, in Italy a ratio of 1 positive patienttested for every 10 infected people is considered reasonable, leading tomore than 1 million infected people (corresponding to more than 1.7% ofthe Italian population) in total.

Identifying not only individuals at risk of infection, but also those athigh risk of morbidity and mortality is one of the most importantchallenges in reducing the social impact of the disease. In addition,the results provided by cellular, genetic and/or biomarker analysescould guide the development and implementation of desperately neededstrategies for the development of new antiviral agents and opportunitiesfor cell therapy.

SUMMARY OF THE INVENTION

The present inventions generally concern methods of identifying subjectsat risk of developing a severe COVID-19 infection and/or severecomplications from COVID-19 infection. For example, in certainembodiments, disclosed herein are methods of identifying a subject atrisk of developing a severe complication from Coronavirus disease 2019(COVID-19) infection, such methods generally comprising a step ofdetermining whether the subject has clonal hematopoiesis ofindeterminate potential (CHIP). In certain aspects, the step ofdetermining whether the subject has CHIP comprises sequencing at leastpart of the genome of one or more cells in a blood sample of the subjectto identify a mutation in one or more genes selected from the groupconsisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS andSF3B1, wherein the presence of said mutation indicates that the subjecthas CHIP and is at risk of developing a severe COVID-19 infection and/ora severe complication from COVID-19 infection.

In certain embodiments, the methods disclosed herein further comprise astep of treating a subject identified as having CHIP, or that isotherwise at risk of developing a severe COVID-19 infection or a severecomplication from COVID-19 infection. For example, in certain aspects ofthe methods disclosed herein treating the subject comprisesadministering to the subject one or more agents or vaccines to reducethe subject's likelihood of contracting COVD-19 infection.

Also described herein are methods of treating a subject at risk ofdeveloping a severe Coronavirus disease 2019 (COVID-19) infection and/orsevere complications from COVID-19 infection. For example, in certainembodiments, disclosed herein are methods of treating a subject at riskof developing a severe COVID-19 infection, such methods generallycomprising a step of determining whether the subject has clonalhematopoiesis of indeterminate potential (CHIP). In certain aspects thestep of determining whether the subject has CHIP comprises sequencing atleast part of the genome of one or more cells in a blood sample of thesubject to identify a mutation in one or more genes selected from thegroup consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRASand SF3B1; wherein the presence of said mutation indicates that thesubject has CHIP and is at risk of developing a severe COVID-19infection; and treating the subject having CHIP to reduce the subject'srisk of contracting a severe COVID-19 infection. For example, in certainaspects of the methods disclosed herein treating the subject comprisesadministering to the subject one or more agents or vaccines to reducethe subject's likelihood of contracting COVD-19 infection.

In certain embodiments of the inventions disclosed herein, the subjectis at risk of requiring hospitalization. In certain embodiments, thesubject is under the age of about 30, 40, 50, 60, 70, 80, 90, 100 yearsold or older. In certain embodiments, the subject has a severeautoimmune disease. In yet other embodiments, the subject has a comorbidcondition (e.g., a comorbid condition selected from the group consistingof cancer, cerebrovascular disease, chronic kidney disease, chronicobstructive pulmonary disease (COPD), diabetes mellitus, heartconditions, obesity, pregnancy, history of smoking, Down syndrome, humanimmunodeficiency virus (HIV), neurologic conditions, interstitial lungdisease, pulmonary fibrosis, pulmonary hypertension, Sickle celldisease, solid organ or blood stem cell transplantation, substance usedisorders, use of corticosteroids or other immunosuppressivemedications, cystic fibrosis, thalassemia, asthma, hypertension, immunedeficiencies, and liver disease). In certain embodiments, the comorbidcondition is selected from the group consisting of cancer,cerebrovascular disease, chronic kidney disease, chronic obstructivepulmonary disease (COPD), diabetes mellitus, heart conditions, obesity,pregnancy, and history of smoking.

In certain aspects, the mutation is a somatic mutation. For example, incertain embodiments of the methods disclosed herein, the mutation is asomatic mutation in DNMT3A and/or TET2.

Also described herein are methods of treating an infection in a subject.The methods may comprise sequencing at least part of the genome of oneor more cells in a blood sample of a subject in need of treatment;identifying in the blood sample a mutation in one or more genes selectedfrom the group consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53,SRSF2, KRAS and SF3B1, wherein the presence of said mutation indicatesan increased susceptibility to an infection; and treating the subject,for example, by vaccinating the subject and/or reducing the incidence ofhematopoietic clones comprising the mutation in the subject's blood.

Also described herein are methods of treating an infection in a subject.The methods may comprise sequencing one or more nucleic acids selectedfrom the group consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53,SRSF2, KRAS, and SF3B1 from one or more cells in a blood sample of thesubject; detecting the presence of a mutation in the sequenced nucleicacids; and treating the subject by reducing the incidence ofhematopoietic clones comprising the mutation (e.g., a mis-sensemutation) in the subject's blood.

The infection may be a viral or a non-viral infection. In someembodiments, the infection is a viral infection and the virus is acoronavirus (e.g., severe acute respiratory syndrome coronavirus(SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV),severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)), influenza(e.g., influenza A (e.g., H1N1, H5N1, H1N2, H2N1, H3N1, H3N2, H2N3) orinfluenza B), or respiratory syncytial virus (RSV). In some embodiments,the infection is a non-viral infection (e.g., a bacterial, fungal,yeast, parasitic, or prion infection). In certain embodiments, theinfection is Coronavirus disease 2019 (COVID-19). In some embodiments,the infection results in respiratory distress and/or a cytokine storm.

Described herein are methods of treating a coronavirus (e.g., severeacute respiratory syndrome coronavirus (SARS-CoV), Middle Eastrespiratory syndrome coronavirus (MERS-CoV), severe acute respiratorysyndrome coronavirus 2 (SARS-CoV-2)) infection in a subject. The methodsmay comprise sequencing at least part of the genome of one or more cellsin a blood sample of a subject in need of treatment; identifying in theblood sample a mutation in one or more genes selected from the groupconsisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS, andSF3B1, wherein the presence of said mutation indicates an increasedsusceptibility to an infection. In certain aspects, the method furthercomprises a step of treating the subject, for example, by vaccinatingthe subject or reducing the incidence of hematopoietic clones comprisingthe mutation in the subject's blood. Alternatively, in certainembodiments, the method further comprises a step of treating the subjectby implementing measures intended to reduce the subject's exposure tothe infection and/or likelihood of contracting such infection (e.g.,self-isolation, quarantine and/or social distancing). In certainaspects, the treatment comprises administering one or more agents orvaccines to the subject to reduce the likelihood of contracting theinfection and/or developing complications therefrom.

Also described herein are methods of predicting a subject'ssusceptibility to a coronavirus infection. The methods may comprisesequencing one or more nucleic acids selected from the group consistingof DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1 fromone or more cells in a blood sample of the subject; and detecting thepresence of a mutation in the sequenced nucleic acids, and wherein thepresence of the mutation indicates an increased susceptibility to acoronavirus infection.

In some embodiments, the gene or nucleic acid is selected from the groupconsisting of TET2, DNMT3A, PPM1D, JAK2, and combinations thereof. Insome embodiments, the mutation is a mis-sense mutation, frame-shiftmutation, nonsense mutation, or splice-site disruption. In someembodiments, the mutation is in DNMT3A in exons 7 to 23. In someembodiments, the mutation is a mis-sense mutation in DNMT3A selectedfrom the group consisting of G543C, S714C, F732C, Y735C, R736C, R749C,F751C, W753C, and L889C. In some embodiments, the mutation is a V617Fmutation in JAK2. In some embodiments, the mutation is a disruptivemutation in TET2. In some embodiments, the mutation is a disruptivemutation in PPM1D.

In some embodiments, the mutation increases inflammation, induces apro-inflammatory state in the subject, or causes elevations ininflammatory cytokines. In some embodiments, the cytokines are orcomprise interleukins (e.g., IL-6, IL-8, IL-1α, IL-1β, IL-12, IL-18),interferons (e.g., IFN-α, IFN-β, IFN-γ, IFN-λ), and/or tumor necrosisfactors (IFN-α, IFN-β, IFN-γ, IFN-λ). In some embodiments, reducing theincidence of hematopoietic clones comprising the mutation in thesubject's blood reduces inflammation or inflammatory cytokines in thesubject.

In some embodiments, the methods described herein further compriseidentifying in the blood sample at least one genetic variant selectedfrom the group consisting of: ACE2, CRP, IL1A, IL1B, IL6, IL6R, CXCL8,IL10, IL12A, IL12B, IL18, IFITM3, TNF, LTA, LTB, IFN-α1, IFN-α2, IFN-α4,IFN-α5, IFN-α6, IFN-α7, IFN-α8, IFN-α10, IFN-α13, IFN-α14, IFN-α16,IFN-α17, IFN-α21, IFN-beta, IFN-epsilon, IFN-gamma, IFN-kappa,IFN-lambda, and IFN-omega.

In some embodiments, the subject is a human subject, and in someembodiments, is at least 20, 30, 40, 50, 60, 70 years of age or older.In some embodiments, the subject has a comorbid condition (e.g.,cardiovascular disease, diabetes, asthma, emphysema, obesity, cancer).

In some embodiments, the one or more cells in the blood sample arenucleated cells. In some embodiments, the one or more cells in the bloodsample are somatic cells.

Also described herein are methods of predicting a subject'ssusceptibility to a coronavirus infection. The methods may comprisesequencing DNMT3A nucleic acids from one or more cells in a blood sampleof a subject; detecting the presence of a mis-sense mutation in thesequenced DNMT3A nucleic acids; sequencing TET2 nucleic acids from oneor more cells in the blood sample of the subject; and detecting thepresence of a disruptive mutation in the sequenced TET2 nucleic acids,wherein the presence of a mis-sense mutation in the sequenced DNMT3Anucleic acids and/or the TET2 nucleic acids indicates the subject'sincreased susceptibility to a coronavirus infection.

Also described herein are methods of treating an infection in a subject.The methods comprise sequencing at least part of the genome of one ormore cells in a blood sample of a subject in need of treatment;identifying in the blood sample a somatic sequence mutation in one ormore genes selected from the group consisting of DNMT3A, TET2, ASXL1,PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1; identifying in the bloodsample a somatic structural chromosomal mutation in one or more genesselected from the group consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2,TP53, SRSF2, KRAS and SF3B1; and/or identifying in the blood sample agermline sequence mutation in one or more genes selected from the groupconsisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS andSF3B1, wherein the presence of one or more somatic sequence mutations,somatic structural mutations or germline sequence mutations indicates anincreased susceptibility to an infection, and treating the subject byreducing the incidence of hematopoietic clones comprising the mutationin the subject's blood.

The above discussed, and many other features and attendant advantages ofthe present inventions will become better understood by reference to thefollowing detailed description of the invention.

DETAILED DESCRIPTION OF THE INVENTION

For reasons that are not entirely clear, some individuals havedysfunctional immune systems that fail to keep a response to Coronavirusdisease 2019 (COVID-19) in check, leading to an uncontrolled immuneresponse. It is generally known that the virus triggers anoverproduction of immune cells and their signaling molecules leading toa cytokine storm often associated with a flood of immune cells into thelungs and ultimately resulting in acute respiratory distress syndrome(ARDS).

Host immune responses to viral infections are generally known tocomprise multiple intricate processes that coordinate together to playsignificant roles in the protection of the host. The understanding ofthe COVID 19 host interaction will require a comprehensive examinationof various cellular, genetic and biomarker factors in a blood sample toelucidate the dynamics of host immune system response, many of which areunclear at this time. Bridging these gaps will pave the way foridentifying not only individuals at risk of being infected but those athigh risk for morbidity and mortality. In addition, the powerful dataprovided by cellular/genetic/biomarker analyses could guide strategiesfor development of novel antiviral agents and opportunities for celltherapy.

Described herein are methods for detecting and/or treating geneticvariations associated with a dysfunctional immune response that resultsin an increased mortality risk in a subject. Also described herein aremethods for identifying and/or treating subjects at risk of, e.g.,susceptible to, a dysfunctional immune response associated with geneticvariations. In some embodiments, a blood sample from the subject issequenced and one or more genetic variations or mutations areidentified. In some embodiments, genetic variations are identified inone or more genes selected from the group consisting of DNMT3A, TET2,ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1. A dysfunctional immuneresponse may include an inflammatory and/or interferon response. In someembodiments, the one or more genetic variations result in increasedmortality risk in a subject having an infection (e.g., COVID-19infection).

Described herein are methods for treating infections in a subjectcomprising sequencing at least part of the genome of one or more cellsin a blood sample of a subject in need of treatment; identifying in theblood sample a mutation in one or more genes selected from the groupconsisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS andSF3B1, wherein the presence of said mutation indicates an increasedsusceptibility to an infection; and treating the subject by reducing theincidence of hematopoietic clones comprising the mutation in thesubject's blood. In some aspects, an identified mutation in the one ormore genes selected from the group consisting of DNMT3A, TET2, ASXL1,PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1 is an indicator of clonalhematopoiesis of indeterminate potential (CHIP).

In some aspects, mutations are somatic sequence mutations, somaticstructural chromosomal mutations, and/or germline sequence mutations. Insome embodiments, somatic sequence mutations in one or more genes areidentified in a blood sample. In some embodiments, somatic structuralchromosomal mutations are identified in a blood sample. In someembodiments, germline sequence mutations are identified in a bloodsample.

Sequencing of DNA can be performed on tissues or cells. Sequencing ofspecific cell types (for example, hematopoietic cells obtained by flowsorting) can identify mutations in specific cell types that providespecific predictive value. Some cell types may provide a greaterpredictive value than other cell types. In some embodiments, the one ormore cells in a blood sample are nuclear cells, e.g., are somatic cells.

Sequencing can also be conducted in single cells, using appropriatesingle-cell sequencing strategies. Single-cell analyses can be used toidentify high-risk combinations of mutations co-occurring in the samecells. Co-occurrence signifies that the mutations are occurring in thesame cell clone and carry a greater risk, and therefore have a greaterpredictive value, than occurrence of the same mutations in differentindividual cells.

In some embodiments, at least part of the genome of one or more cells ina blood sample, e.g., of a subject in need of treatment, is sequenced.In some embodiments the part of the genome that is sequenced is limitedto specific genes, the whole exome, or parts of an exome. For example,the sequencing may be whole exome sequencing (WES). Sequencing can becarried out according to any suitable technique, many of which aregenerally known in the art. Many proprietary sequencing systems areavailable commercially and can be used in the context of the presentinvention, such as for example from Illumina, USA. Single-cellsequencing methods are known in the art, as noted for example byEberwine et al., Nature Methods 11, 25-27 (2014) doi:10.1038/nmeth.2769Published online 30 Dec. 2013; and especially single cell sequencing inmicrofluidic droplets (Nature 510, 363-369 (2014)doi:10.1038/nature13437), the entire contents of which are incorporatedherein by reference.

Sequencing may be performed of specific genes only, specific parts ofthe genome, or the whole genome. In some aspects, specific parts of agene can be sequenced; for example, in DNMT3A, exons 7 to 23 can besequenced. Where a part of a genome is sequenced, that part can be theexome. The exome is the part of the genome formed by exons, and thus anexon sequencing method sequences the expressed sequences in the genome.There are 180,000 exons in the human genome, which constitute about 1%of the genome, or approximately 30 million base pairs. Exome sequencingrequires enrichment of sequencing targets for exome sequences; severaltechniques can be used, including PCR, molecular inversion probes,hybrid capture of targets, and solution capture of targets. Sequencingof targets can be conducted by any suitable technique.

Methods of identifying somatic structural chromosomal mutations andgermline sequence mutations in blood samples are known to those of skillin the art. Examples of such methods are described in WO 2019/079493 andUS 2017/0321284, both of which are incorporated herein by reference.

In some embodiments, a mutation (e.g., a mutation in one or more genesselected from the group consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2,TP53, SRSF2, KRAS and SF3B1) is identified in a blood sample (e.g., ablood sample from a subject). In certain aspects, the presence of amutation indicates an increased susceptibility to an infection (e.g., ahuman subject's increased susceptibility to COVID-19 infection and/orthe corresponding sequalae). In some aspects, the presence of a mutationindicates an increased susceptibility to an adverse clinical outcome dueto a dysfunctional immune response (e.g., a human subject's increasedsusceptibility to developing respiratory distress, ARDS and/or cytokinestorm). In some embodiments, a mutation in one or more genes is selectedfrom the group consisting of DNMT3A, TET2, PPM1D, and JAK2. In certainembodiments, a mutation in DNMT3A is identified, e.g., in a bloodsample. In certain embodiments, a mutation in TET2 is identified, e.g.,in a blood sample. In certain embodiments, a mutation in PPM1D isidentified, e.g., in a blood sample. In certain embodiments, a mutationin JAK2 is identified, e.g., in a blood sample.

DNMT3A is DNA (cytosine-5-1-methyltransferase 3 alpha and is encoded onchromosome 2 (HGMC 2978). ASXL1 is additional sex combs liketranscriptional regulator 1 and is encoded on chromosome 20 (HGNC18318). TET2 is tet methylcytosine dioxygenase 2 and is encoded onchromosome 4 (HGNC 25941). PPM1D is protein phosphatase, Mg2+/Mn2+dependent, 1D and is encoded on chromosome 17 (HGNC 9277). JAK2 is januskinase 2 and is encoded on chromosome 9 (HGNC 6192). TP53 is tumorprotein p53 and is encoded on chromosome 17 (HGNC 11998). SRSF2 isserine and arginine rich splicing factor 2 and is encoded on chromosome17 (HGNC 10783). KRAS is KRAS proto-oncogene and is encoded onchromosome 12 (HGNC 6407). SF3B1 is splicing factor 3b subunit 1 and isencoded on chromosome 2 (HGNC 10768).

Mutations in genes can be disruptive (e.g., they have an observed orpredicted effect on protein function) or non-disruptive. Anon-disruptive mutation is typically a mis-sense mutation, in which acodon is altered such that it codes for a different amino acid, but theencoded protein is still expressed. In some embodiments, somaticmutations may be mis-sense mutations or disruptive mutations (e.g.,frame-shift, nonsense, or splice-site disruptions).

Putative somatic mutations include but are not limited to those allelesthat comprise at least one of non-silent/disruptive nucleotide changes,indels, mis-sense mutations, frameshifts, stop mutations (addition ordeletion), read-through mutations, splice mutations; and a confirmedchange not due to a sequencing error or artifact of the testing system.

In some embodiments, mutations in DNMT3A are predominantly mis-sensemutations. In some aspects, mutations (e.g., mis-sense mutations) inDNMT3A are localized in exons 7 to 23. In some aspects, mutations inDNMT3A are enriched for cysteine-forming mutations. A common base-pairchange in somatic variants is a cytosine-to-thymine transition. In someembodiments, a mutation in DNMT3A is a mis-sense mutation selected fromthe group consisting of G543C, S714C, F732C, Y735C, R736C, R749C, F751C,W753C, and L889C. In some embodiments, mutations in TET2 and/or PPM1Dare disruptive mutations. In some embodiments, a mutation in JAK2 is aV617F mutation. Additional non-limiting examples of mutations found inDNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1 aredescribed in: Genovese, et al., “Clonal Hematopoiesis and Blood-CancerRisk Inferred from Blood DNA Sequence” N Engl J Med (2014)371:2477-2487; Jaisawal et al., “Age-Related Clonal HematopoiesisAssociated with Adverse Outcomes” N Engl J Med (2014) 371:2488-2498;Bick, et al. “Genetic Interleukin 6 Signaling Deficiency AttenuatesCardiovascular Risk in Clonal Hematopoiesis” Circulation (2020)141:124-131; Cook et al., “Clonal hematopoiesis and inflammation:Partners in leukemogenesis and comorbidity” Exp Hematol (2020) 83:85-94;Perner et al., “Roles of JAK2 in Aging, Inflammation, Hematopoiesis andMalignant Transformation” Cells (2019) 8(8) 854; Cook et al., “Comorbidand inflammatory characteristics of genetic subtypes of clonalhematopoiesis” Blood Adv. (2019) 3(16): 2482-2486; Uyanik et al., “DNAdamage-induced phosphatase Wip1 in regulation of hematopoiesis, immunesystem and inflammation” Cell Death Discov (2017) 3, 17018; US2017/0321284; and WO 2019/079493, all incorporated herein by reference.

In some embodiments, a blood sample obtained from a subject may furtherbe sequenced to identify at least one additional genetic variant. Forexample, a blood sample may be sequenced to identify at least onegenetic variant for a tumor necrosis factor (e.g., TNF, LTA, and LTB),interferon (e.g., IFN-α1, IFN-α2, IFN-α4, IFN-α5, IFN-α6, IFN-α7,IFN-α8, IFN-α10, IFN-α13, IFN-α14, IFN-α16, IFN-α17, IFN-α21, IFN-beta,IFN-epsilon, IFN-gamma, IFN-kappa, IFN-lambda, and IFN-omega), orinterleukin (e.g., IL1A, IL1B, IL6, IL6R, CXCL8, IL10, IL12A, IL12B, andIL18). In some embodiments, at least one genetic variant is selectedfrom the group consisting of ACE2, CRP, IL1A, IL1B, IL6, IL6R, CXCL8,IL10, IL12A, IL12B, IL18, IFITM3, TNF, LTA, LTB, IFN-α1, IFN-α2, IFN-α4,IFN-α5, IFN-α6, IFN-α7, IFN-α8, IFN-α10, IFN-α13, IFN-α14, IFN-α16,IFN-α17, IFN-α21, IFN-beta, IFN-epsilon, IFN-gamma, IFN-kappa,IFN-lambda, and IFN-omega.

A mutation in one or more genes may cause an increase in inflammation,induce a pro-inflammatory state in a subject, or cause an elevation ininflammatory cytokines (e.g., interleukins, interferons, and/or tumornecrosis factors). Examples of inflammatory cytokines include IL-6,IL-8, IL-1α, IL-1β, IL-12, IL-18, IFN-α, IFN-β, IFN-γ, IFN-λ, TNF-α,TNF-β, and TNF-γ.

In some embodiments, a subject has or is susceptible to an infection. Aninfection may be a viral infection or a non-viral infection. Forexample, a viral infection may be caused by a coronavirus, influenza,respiratory syncytial virus (RSV), among others. In some embodiments, acoronavirus is selected from the group consisting of severe acuterespiratory syndrome coronavirus (SARS-CoV), Middle East respiratorysyndrome coronavirus (MERS-CoV), and severe acute respiratory syndromecoronavirus 2 (SARS-CoV-2). In certain embodiments, an infection isCoronavirus disease 2019 (COVID-19). In some embodiments, influenza isinfluenza A or influenza B. Strains of influenza A include H1N1, H5N1,H1N2, H2N1, H3N1, H3N2, and H2N3. In some embodiments, a non-viralinfection is a bacterial, yeast (e.g., fungal), parasitic, or prioninfection. In some aspects, the viral or non-viral infection results inrespiratory distress or complications (e.g., acute respiratory distresssyndrome, interstitial pneumonia, other respiratory disease) and/or acytokine storm. Additional symptoms or complications of the infectioninclude lung damage, blood complications (e.g., blood clots, hemostaticderangement), central nervous system complications (e.g., encephalitis),kidney damage, organ failure, acute cardiac injury (e.g., cardiacarrest, coronary artery occlusion, heart failure, myocarditis,cardiomyopathy), and loss of smell or taste.

In some embodiments, a subject may be treated by reducing the incidenceof hematopoietic clones comprising a mutation in one or more genes inthe subject's blood. “Treat, “treatment,” “treated,” “treating,” etc.refer to providing medical or surgical attention, care, or management toan individual. The individual is usually ill or injured, or at increasedrisk of becoming ill relative to an average member of the population andin need of such attention, care, or management. Treating can refer toprolonging survival as compared to expected survival if not receivingtreatment. Thus, one of skill in the art realizes that a treatment mayimprove the disease condition, but may not be a complete cure for thedisease. As used herein, the term “treatment” includes prophylaxis, andmay further include the implementation of measures intended to reducethe subject's likelihood of contracting an infectious disease (e.g.,COVID-19). For example, in certain aspects, the treatment may includemedical and/or pharmacologic interventions intended to reduce thesubject's likelihood of contracting an infectious disease (e.g., theprophylactic administration of one or more vaccines, convalescent plasmainfusion and/or antibodies to the subject). In other embodiments, thetreatment may include non-pharmacologic interventions, such asself-isolation and/or quarantine of a subject identified as beingsusceptible to developing an infectious disease and/or the associatedcomplications. Alternatively, treatment is “effective” if the disease isprevented and/or if progression of a disease is reduced or halted.“Treatment” can also mean prolonging survival as compared to expectedsurvival if not receiving treatment.

A treatment is not necessarily curative, and in certain aspects mayreduce the effect of an infection by a certain percentage over anuntreated infection. For example, treatment may reduce inflammation orreduce the presence of inflammatory cytokines in a subject sufferingfrom an infection. In some embodiments, methods of treatment may bepersonalized medicine procedures, in which the DNA of an individual isanalyzed to provide guidance on the appropriate therapy for thatspecific individual. The methods of the invention may provide guidanceas to whether treatment is necessary or most effective, as well asrevealing progress of the treatment and guiding the requirement forfurther treatment of the individual.

It should be noted that in certain aspects, the treatment may comprisethe recommendation and/or implementation of measures intended to reducethe subject's likelihood of contracting an infection, and which measuresare not isolated to the subject. For example, in certain embodiments,the treatment may comprise vaccinating members of the subject'scommunity (e.g., family, co-workers and/or healthcare providers) in aneffort to reduce the transmission of the infection.

In some embodiments, treatment comprises reducing the incidence of thepresence of clonal hematopoiesis in the subject's blood. In otherembodiments, treatment or monitoring includes repeating the sequencingof a blood sample of the subject monthly, bi-monthly, or quarterly, andreducing (or monitoring reduction in) the incidence of the presence ofclonal hematopoiesis in the subject's blood. In some embodiments,treatment or monitoring comprises including the subject as a candidateto receive a bone marrow transplant. In some embodiments, treatmentincludes administering to the subject a bone marrow transplant. In someembodiments, treatment includes transfusing the subject with blood inwhich clonal hematopoiesis is absent.

In some aspects, a subject is identified as being high risk based ongene sequencing (e.g., performing a CHIP assessment). A subjectidentified as being high risk has an increased risk of contracting aninfection and/or of having a serious reaction to an infection. In someaspects, high risk subjects are treated by early targeting ofinflammation. For example, a high risk subject may be treated byadministering pro-inflammatory driver inhibitors and/or low-dosehypomethylating drugs. In some embodiments, high risk subjects may betreated by vaccinating the subject and/or vaccinating members of thesubject's community to reduce the transmission of the infection, andthereby reduce the likelihood that the subject will become infected.

In some aspects, vaccinating the subject and/or vaccinating members ofthe subject's community to reduce the transmission of the infectioncomprising the administration of a single vaccine dose or theadministration of two vaccine doses. One or more vaccine boosters mayoptionally be administered after administration of the initial vaccinedose(s). For example, a vaccine booster may be administered about 6months, 9 months, 12 months, 15 months, 18 months, 21 months, or 24months after receiving the initial vaccine dose(s). In some aspects, avaccine booster may be administered yearly. In some embodiments, thevaccine is an mRNA vaccine or a viral vector vaccine. In one embodiment,the vaccine is a BNT162b2 vaccine (a lipid nanoparticle-formulated,5nucleoside-modified RNA (modRNA) encoding the SARS-CoV-2 full-lengthspike, modified by two proline mutations to lock it in the prefusionconformation). In one embodiment, the vaccine is a mRNA-1273 vaccine (alipid nanoparticle-encapsulated mRNA-based vaccine that encodes theprefusion stabilized full-length spike protein of the severe acuterespiratory syndrome coronavirus 2 (SARS-CoV-2)). In one embodiment, thevaccine is a Ad26.COV2.S Vaccine (a monovalent vaccine composed of arecombinant, replication-incompetent human adenovirus type 26 (Ad26)vector that encodes a SARS-CoV-2 spike (S) protein in a stabilizedconformation). In one embodiment, the vaccine is a ChAdOx1 nCoV-19vaccine (a replication-deficient chimpanzee adenoviral vector ChAdOx1,containing the SARS-CoV-2 structural surface glycoprotein antigen (spikeprotein; nCoV-19) gene).

In some aspects, treating a subject for an infection includes adjuvanttreatment. For example, adjuvant treatment for preventing sequelae ofacute pulmonary disease may include administration of one or moreantiviral medications (e.g., remdesivir), anti-inflammatory medications,antibiotics, plasmapheresis, convalescent plasma infusion, monoclonalantibodies (e.g., bamlanivimab, etesevimab, casirivimab, imdevimab,recombinant humanized anti-interleukin-6 receptor monoclonal antibodies(e.g., tocilizumab)), corticosteroids (e.g., dexamethasone),supplemental oxygen, and mesenchymal stem cell/iPS cell therapy. In someaspects, adjuvant treatment for preventing sequelae of cardiac and/orinfectious disease includes administration of one or more antiviralmedications, anti-inflammatory medications, antibiotics, plasmapheresis,convalescent plasma, mesenchymal stem cell/iPS cell therapy, bloodthinners, anti-arrhythmic drugs, and cardioprotective drugs. In certainaspects, blood pressure medications may be administered to a subjectincluding angiotensin converting enzyme (ACE) inhibitors and/orangiotensin receptor blockers (ARBs).

In some embodiments, a subject's susceptibility to an infection ispredicted using the sequencing and detection methods described herein.For example, a subject's susceptibility to an infection (e.g., acoronavirus infection) may be predicted by sequencing one or morenucleic acids selected from the group consisting of DNMT3A, TET2, ASXL1,PPM1D, JAK2, TP53, SRSF2, KRAS and SF3B1 from one or more cells in ablood sample of the subject; and detecting the presence of a mutation inthe sequenced nucleic acids. In some embodiments, the presence of themutation indicates an increased susceptibility to a coronavirus (e.g.,COVID-19) infection. In some embodiments, a subject's increasedmortality risk, e.g., resulting from a dysfunctional immune response, ispredicted using the sequencing and detection methods described herein.In some aspects, the dysfunctional immune response is an inflammatory orinterferon response.

In certain embodiments, a subject's susceptibility to an infection(e.g., a coronavirus infection) is predicted by sequencing DNMT3A andTET2 nucleic acids from one or more cells in a blood sample from thesubject and detecting the presence of a mutation (e.g., mis-sensemutations or disruptive mutations) in the sequenced nucleic acids. Thepresence of a mutation in the sequenced DNMT3A and/or TET2 nucleic acidsmay indicate a subject's increased susceptibility to the infection.

In certain embodiments, the subject is a mammal, e.g., a primate, e.g.,a human. The terms, “patient” and “subject” are used interchangeablyherein. Preferably, the subject is a mammal. The mammal can be a human,non-human primate, mouse, rat, dog, cat, horse, or cow, but are notlimited to these examples. In certain embodiments, the subject is ahuman.

The subject may be any age from birth to death, e.g., the subject may bean infant, a toddler, a child, a teenager, a young adult, or an adult.The subject may be at least about 1 day old, 1 year old, 5 years old, 10years old, 20 years old, 30 years old, 40 years old, 50 years old, 60years old, 70 years old, 80 years old, 90 years old, or 100 years old.In some embodiments, the subject is under the age of 40, between theages of 40-60, or over the age of 60.

In other embodiments, the subject may exhibit one or more risk factorsincluding asthma, chronic lung disease, interstitial lung disease,cystic fibrosis, pulmonary hypertension, chronic obstructive pulmonarydisease (COPD), diabetes, obesity, being aged 65 years or older, heartcondition (e.g., heart failure, coronary artery disease,cardiomyopathies, hypertension, etc.), high blood pressure, chronickidney disease, immunocompromised (e.g., undergoing therapy for cancer),liver disease, dementia or other neurological condition, down syndrome,HIV infection, pregnancy, sickle cell disease or thalassemia, stroke orcerebrovascular disease, solid organ or blood stem cell transplant,substance use disorder, or history of being a smoker. In someembodiments, the subject has a comorbid condition selected from thegroup consisting of cardiovascular disease, lung disease, kidneydisease, liver disease, neurological condition, cerebrovascular disease,diabetes, obesity, asthma, emphysema, and cancer.

It is to be understood that the invention is not limited in itsapplication to the details set forth in the description or asexemplified. The invention encompasses other embodiments and is capableof being practiced or carried out in various ways. Also, it is to beunderstood that the phraseology and terminology employed herein is forthe purpose of description and should not be regarded as limiting.

While certain compounds, compositions and methods of the presentinvention have been described with specificity in accordance withcertain embodiments, the following examples serve only to illustrate themethods and compositions of the invention and are not intended to limitthe same.

The articles “a” and “an” as used herein in the specification and in theclaims, unless clearly indicated to the contrary, should be understoodto include the plural referents. Claims or descriptions that include“or” between one or more members of a group are considered satisfied ifone, more than one, or all of the group members are present in, employedin, or otherwise relevant to a given product or process unless indicatedto the contrary or otherwise evident from the context. The inventionincludes embodiments in which exactly one member of the group is presentin, employed in, or otherwise relevant to a given product or process.The invention also includes embodiments in which more than one, or theentire group members are present in, employed in, or otherwise relevantto a given product or process. Furthermore, it is to be understood thatthe invention encompasses all variations, combinations, and permutationsin which one or more limitations, elements, clauses, descriptive terms,etc., from one or more of the listed claims is introduced into anotherclaim dependent on the same base claim (or, as relevant, any otherclaim) unless otherwise indicated or unless it would be evident to oneof ordinary skill in the art that a contradiction or inconsistency wouldarise. Where elements are presented as lists, (e.g., in Markush group orsimilar format) it is to be understood that each subgroup of theelements is also disclosed, and any element(s) can be removed from thegroup. It should be understood that, in general, where the invention, oraspects of the invention, is/are referred to as comprising particularelements, features, etc., certain embodiments of the invention oraspects of the invention consist, or consist essentially of, suchelements, features, etc. For purposes of simplicity those embodimentshave not in every case been specifically set forth in so many wordsherein. It should also be understood that any embodiment or aspect ofthe invention can be explicitly excluded from the claims, regardless ofwhether the specific exclusion is recited in the specification. Thepublications and other reference materials referenced herein to describethe background of the invention and to provide additional detailregarding its practice are hereby incorporated by reference.

EXEMPLIFICATION Example 1

A DNA microarray will be used to identify, and report inherited geneticvariants (e.g., TET2, DNMT3A, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS,and/or SF3B1) associated with a variety of cardiovascular and metabolicdisorders, as well as with certain cancers. These cancers and disorders,if pre-existing, can increase the risk of death related to COVID 19complications. Also included in this microarray are genetic variantsassociated with pro- and anti-inflammatory effects. Some of theseinclude ACE2, CRP, IL-6R, IL-1A GC, and IL-10. Furthermore, themicroarray may elucidate which inborn errors of immunity can causelife-threatening COVID 19 in previously healthy younger patients.

In addition, the microarray may test for various biomarkers present inthe blood that are involved in immunity and inflammation, which mayimpact COVID 19 infection response. These biomarkers may include hsCRP,IL-6, IL-8, and TNF-alpha.

Clonal hematopoiesis of indeterminate potential (CHIP) is a geneticissue unique to nucleated cells contained within the blood. CHIP refersto the unhealthy expansion of the same set of detrimental mutations anddamage to an individual's DNA in their nucleated blood cells. CHIP maypotentially explain the age-related prevalence of inflammatoryconditions. A connection between CHIP mutations and heart diseases isbelieved to be mediated by inflammation. It has been more recentlypostulated that CHIP may be associated with broader biological impactwhich may be explained by the fact that people with CHIP have anaberrant inflammatory system compared with those without CHIP.Specifically, recent research has demonstrated that structural DNAdamage in genes such as TET2 and DNMT3A is associated with adysfunctional response to inflammation and infection. In addition, keycytokines such as serum IL-6 and IL-8 have been shown to be elevated inpeople with CHIP, establishing its relevance to the human systemicinflammatory landscape and its impact on poorer health status, andincreased and potentially novel comorbidities.

It is known that somatic exonic mutations are acquired at a rate ofapproximately 1.3 per hematopoietic stem cell per decade and are rarelydetected in people under age 40 years. Clonal hematopoiesis increaseswith age and is found in approximately 10 percent of people withoutknown hematologic malignancies older than 65 years, in almost 12 percentof those aged 80 to 89 years, and in more than 18 percent of those olderthan 90 years.

In addition to detecting and monitoring somatic changes associated withCHIP, a screen will also be performed for germline/inherited variantsassociated with cardiovascular disease, cancer, and inflammation. Theassociation between CHIP, inflammation, and leukocyte dysfunction willbe examined so the CHIP-related comorbidities in COVID 19 can be moreeffectively managed clinically. For example, the presence of CHIP as adisease modifier might explain why some patients are more responsive orrefractory to treatments, or why some patients undergo more slow orrapid disease progression. Furthermore, this dynamic measurement mayelucidate novel opportunities to develop new drug therapies focusing onthe genetic abnormalities of CHIP.

Within the PBMCs that are isolated and preserved from the peripheralblood, a variety of potentially therapeutic cell types will be accessed,including small numbers of CD34+ hematopoietic stem cells (HSCs),T-cells and circulating endothelial progenitor cells (EPCs) that allowfor the derivation of clinical grade induced pluripotent stem (iPS)cells. Notably iPS cells can be programmed to produce virtually any celltype in the human body, such as immune cells and alveolar cells of thelung which produce surfactant (AE2). AE2 cells play various roles inalveolar fluid balance, coagulation/fibrinolysis, and host defense. AE2cells proliferate, differentiate into AE1 cells, and contribute toepithelial repair and immunoregulation. In addition, iPS cells can bedifferentiated into mesenchymal stem cells (MSCs) which are known tomodulate the pro-inflammatory immune cytokine response which is largelyresponsible for the cascade of events leading to severe pulmonarydisease. Whether these cells can restore healthy lung tissue after aCOVID 19 infection will be examined. Blood-derived iPS cells represent apersonalized cell resource for individuals to generate AE2 cells andcells of immune function as a therapy option.

We will provide genetic evaluation for a better understanding of thegenetic history and at risk disease potential, tracking of biomarkers toevaluate progression of disease, measurement of somatic damage andmutations that may contribute to a deranged inflammatory response, andthe opportunity to utilize stem cells in potential therapies,potentially including those for COVID 19 or other viral disease.

Example 2

CHIP is a genetic issue unique to the nucleated cells contained withinblood. It refers to the unhealthy expansion of the same set ofdetrimental mutations and damage to an individual's DNA in theirnucleated blood cells. CHIP may potentially explain the age-relatedprevalence of inflammatory conditions. A connection between CHIPmutations and heart diseases is believed to be mediated by inflammation.It has been more recently postulated that CHIP may be associated withbroader biological impact which may be explained by the fact that peoplewith CHIP have an aberrant inflammatory system compared with thosewithout CHIP. Specifically, key cytokines such as serum IL-6 and IL-8have been shown to be elevated in people with CHIP, establishing itsrelevance to the human systemic inflammatory landscape and its impact onpoorer health status, and increased and potentially novel comorbidities.

It is known that somatic exonic mutations are acquired at a rate ofapproximately 1.3 per hematopoietic stem cell per decade, and are rarelydetected in people under age 40 years. Clonal hematopoiesis increaseswith age and is found in approximately 10 percent of people withoutknown hematologic malignancies older than 65 years, in almost 12 percentof those aged 80 to 89 years, and in more than 18 percent of those olderthan 90 years.

Whether an age-related increase in CHIP contributes to the age-relatedrisk of COVID 19 will be assessed. If these phenomena are shown to belinked, the CHIP-related analysis will provide a quantifiable riskassessment. If the complex association between CHIP, inflammation, andleukocyte dysfunction can be unraveled, CHIP-related comorbidities inCOVID 19 may be more effectively managed clinically. For example, thepresence of CHIP as a disease modifier may explain why some patients aremore responsive or refractory to treatments, or why some patientsundergo more slow or rapid disease progression.

By monitoring acquired changes to both sequence and structure of the DNAwithin the blood it may ultimately provide an additional layer ofinsight into identifying those at increased risk for infection or COVID19 complicated mortality. Somatic changes associated with CHIP may bedetected and monitored, as well as screening for germline/inheritedvariants associated with cardiovascular disease, cancer, andinflammation. Blood samples will be evaluated with a target sequencepanel specific to genes with somatic (accumulated) and germline(inherited) variants associated with CHIP, including but not limited toTET2 and DNMT3A, both of which have been associated with a dysfunctionalresponse to inflammation and infection. In addition, microarray-basedmethods are used to measure somatic structural DNA variation and damageacross the genome, additionally associated with CHIP. Genetic testresults will be correlated with serologic and clinical data from thesubject's medical record.

Example 3

Blood samples will be collected from a variety of subjects. The bloodsamples will be assessed to identify samples that are seropositive COVID19, Influenza A, Influenza B, RSV, SARS-CoV, and MERS-CoV. In addition,normal control blood samples will also be obtained. The blood sampleswill be assayed for clonal hematopoiesis (CHIP) using an assay thatscreens for somatic variations in TET2, DNMT3A, ASXL1, PPM1D, JAK2,TP53, SRSF2, KRAS, and SF3B1.

Additional assessments will be performed on the blood samples includingevaluating the samples for additional variants in other genes, as wellas performing additional blood analysis. Further, clinical data will becollected for the subjects from who the blood samples were taken from.

In addition, the samples will be evaluated for one or more variants inACE2, IFITM3, IL-1A, IL-1B, IL-6, IL-6R, CXCL8, IL-12A, IL-12B, IL-18,TNF-α, TNF-β, and TNF-γ. The samples will be assessed for additionalfeatures such as arterial blood gas, white count with differential(lymphocytes, neutrophils, cytokine levels, C-reactive protein (CRP),erythrocyte sedimentation rate (ESR), Lactase, MB-CK enzyme, etc.)Clinical data for the subjects from whom the blood samples are obtainedwill additionally be considered including, but not limited to, thedevelopment of any complications related to the diagnosis (MI,myocarditis, pulmonary embolism), vital signs, O₂ sat, O₂ requirements,intubation status, mortality, and cause(s) of death. Additionally, anyimaging obtained from the subject will be considered, including chest Xrays (CXR) and CT scans. Finally, any co-morbidities and family historywill also be taken into consideration.

CHIP may be utilized as a disease modifier to explain why some COVID 19patients undergo a faster and more severe disease progression, and whysome patients are more responsive/refractory to treatments. In addition,CHIP may be utilized as a disease modifier in determining which patientsare more susceptible to poor cardiac outcomes. CHIP may also be utilizedto provide guidance on mitigation strategies for population management,e.g., establishing quarantine protocols and identifying groups ofindividuals to be quarantined, possibly more restrictively than otherindividuals.

Example 4 Introduction

The arrival of the newly discovered coronavirus in 2019 has impacted theworld gravely, with over 136 million reported cases and over 3 milliondeaths worldwide. To date, no effective intervention strategies are inplace, yet the emergence of mutant strains is dispersing rapidly. Inaddition, molecular rRT-PCR tests may result in false-negative orfalse-positive outcomes thereby questioning its status as ‘goldenstandard’ for laboratory diagnosis of coronavirus disease 2019(COVID-19) (1). Novel effective diagnostic solutions could be the key tocurbing the spread of COVID-19, as early diagnosis is crucial forcontrolling infectious propagation.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-COV-2) is anenveloped single-stranded, positive-sense ribonucleic acid (ssRNA+)coronavirus which belongs to the family of Coronaviridae (2). Theinfection manifests itself as mild-moderate symptoms, such as coughing,muscle fatigue and fever to severe respiratory failure which requiresICU care and ventilation (3).

Important predictors of severe clinical outcomes are advanced age,gender and comorbid conditions including obesity, diabetes,cardiovascular diseases (CVDs) and chronic obstructive pulmonary disease(COPD) (4-7). Furthermore, emerging data indicate that uncontrolledinflammation is associated with disease severity. Indeed, unfetteredrelease of pro-inflammatory cytokines such as IL-6, IL-2, IL-7 andTNF-alpha can cause a so-called cytokine storm, a potentiallife-threatening phenomenon frequently seen in COVID-19 patients withsevere infection (8). In these patients, the hyperactive immuneactivation may progress to acute respiratory stress syndrome (ARDS) andmultiple organ dysfunctions (9). The presence of a cytokine storm maycontribute to cardiovascular complications, including unstable plaqueformation, coronary artery occlusion, myocardial infarction and,eventually, transition to heart failure (10, 11).

While it is evident that inflammation and pre-existing comorbiditiesplay a significant role in the pathogenesis of COVID-19, the presence ofsomatic acquired mutations has now been recognized as possibly anotherindependent determinant for disease outcome (12, 26). These mutationsare found in genes involved in epigenetic regulation including DNMT3A,TET2, ASXL1 and JAK2, and result in expansion of genetically identicalclones of nucleated peripheral blood stem and progenitor cells. Thisphenomenon is referred to as clonal hematopoiesis of indeterminatepotential (CHIP) and affects a large proportion (>10%) of the olderpopulation over the age of 70 (13). Individuals with CHIP are at anincreased risk for developing CVDs, including aortic valve stenosis,venous thrombosis, and heart failure (14-16). Growing evidence indicatesthat the heightened risk might be due to hyper-inflammatory potential inthese individuals (17-19). Indeed, a TET2 deletion has been associatedwith inflammasome activation and subsequent excessive levels ofpro-inflammatory cytokines IL-6 and IL-1, whereas DNMT3A mutations haveshown increased numbers of chemokines and reduced type-1-interferonsecretion compared to non-CHIP carriers (20-23). Remarkably, however,individuals with large DNMT3A or TET2 CHIP-driver mutations (allelefraction of >10%) were protected from cardiovascular events if they alsohad a simultaneous germline sequence variation in the IL-6 receptor(IL-6R) (24). While most of these outcomes were studied in monocytes andmacrophages, CHIP mutations have also been shown to alter the functionof other leukocyte populations (25). Thus, the presence of CHIP mightdysregulate the entire inflammatory system and therefore may corruptdisease outcome in COVID-19 patients.

In support of this notion, recent clinical data showed higher CHIPfrequencies in patients with severe COVID-19 compared to the controlpopulation (26-27). It should be noted, however, that the control groupof one study, referenced herein as the Duployez study (26) consistedmostly of individuals with unexplained cytopenia, thereby potentiallyintroducing biases that underestimate the overall risk of CHIP.Furthermore, Shivarov, et al. consolidated data of four large studies onthe frequency of CHIP and observed a linear correlation between thepresence of CHIP clones and mortality rate in COVID-19-infected patients(12). This suggests that detection of CHIP may be predictive of fataloutcome in patients with COVID-19. However, to date no clinical studieshave been identified that have conducted a comparison of CHIP prevalenceto mild and severe COVID-19 symptoms.

In the present study, the hypothesis that CHIP is associated withdisease severity in COVID-19 patients was tested. Three age groups wereevaluated—under 40 years-old, 40-60 years old and those older than 65years. While CHIP has been reported to be rare in younger people withfrequencies of about 1 in 1000 in those younger than 40 and about 0.7%in those under 50, CHIP has been more recently found in higherfrequencies in younger individuals with early-onset myocardialinfarction and in some advanced autoimmune disease, and hence it washypothesized that it could be a risk factor in younger populations withsevere COVID-19 disease (27, 29).

Methods

Patients and Samples—Patient samples were collected both throughcollaboration and under the IRB of the New York Blood Center (NYBC) aswell as via a prospective procurement contract through iSpecimen.Patients included in the study were confirmed to be PCR positive forSARS-CoV-2 infection and classified as either severe hospitalization ornon-hospitalized. Severe hospitalized patients were defined as infectedand admitted to the hospital for a minimum of seven days, with apriority towards patients admitted to the ICU and requiring oxygensupport. Non-hospitalized patients were defined as infected andsymptomatic (fever, loss of smell, body aches, coughing) orasymptomatic, but not requiring hospitalization. Samples were excludedfrom collection and/or analysis if the patient had previously receivedblood stem cell transplant or had undergone chemotherapy. Samplesprocured through NYBC were delivered as cryopreserved PBMCs whilesamples received from iSpecimen were supplied as frozen whole blood(WB).

Molecular Analysis—Genomic DNA (gDNA) was extracted from cryopreservedPBMC and frozen WB samples using the PerkinElmer Chemagic 360 B5Kautomated extraction platform. For targeted sequencing extracted gDNAwas normalized and aliquoted to 25 ng/ul in 25 ul total volume. CustomDNA target sequencing libraries were constructed using AgilentSureSelect chemistry. The resulting libraries were analyzed on theAgilent 4200 TapeStation System and quantified by KAPA qPCR. Sampleswere sequenced using the Illumina MiSeq platform with an averagessequencing coverage>1000× across amplicons. DNA sequence quality wasverified and then BAM (Binary Alignment/Map) and VCF (Variant CallFormat) files were generated. Putative somatic variants were annotatedagainst GenBank gene models and the ClinVar database. Both somatic andgermline coding variants from genes DNMT3A, TET2, ASXL1, PPM1D, JAK2,TP53, SF3B1, SRSF2, and KRAS were extracted from the annotated VCF. Uponanalysis CHIP accumulated/somatic variants were defined as having >1.5%variant allele frequency (VAF) and 20 or more variant cells.

For microarray analysis, extracted gDNA was normalized and aliquoted to25 ng/ul in 50 ul total volume. Normalized and aliquoted gDNA sampleswere run on the Illumina Infinium™ Global Screening Array (GSA)—24 v2.0BeadChip in the labs of Diagnomics (San Diego, Calif. USA). Theresulting iDAT files were converted to GTC files using the IlluminaArray Analysis Platform and a VCF file with genotype information wasgenerated. SNP and indel markers were annotated against GenBank genemodels and a table with all the genotyped markers from genes IL6R,CXCL8, IL6, and ACE2 were extracted from the VCF using theBCFtools+split−vep plugin.

Results

The study described herein assessed whether CHIP is associated withdisease severity in COVID-19 patients by looking at three age groups ofpatients—under 40 years-old, 40-60 years old and older than 65 years.The most common somatic mutations identified were DNMT3A and TET2 in allgroups, with 54% in those <60, and 71% in those over the age of 65. Asummary of the individual groups is provided below.

Age Group Under 40

The cohort of severe disease in the under 40 group included 9 patients.Two patients (22.2%) had CHIP with VAF>1.5%. One of these patients hadtwo CHIP mutations (DNMT3A and SF3B1) both with a VAF>1.5%. A thirdpatient had a somatic mutation in TP53 with a VAF of 0.8% overallresulting in 3 patients (33.3%) having detectable somatic CHIPmutations. The expected CHIP in individuals under the age of 40 is about0.1%. In addition, a fourth patient in this group had a germlinemutation in TP53. Collection of samples from COVID-19 infected butnon-hospitalized individuals in this age group is under way.

Age Group 40-60

The cohort in the 40-60 age group included 36 patients with severedisease and 4 non-hospitalized infected patients. Three (8.3%) of the 36patients with severe disease had CHIP with VAF>1.5% with the followingmutations: TET2, DNMT3A and KRAS. In addition, 7 additional patientswith severe disease had CHIP mutations (3 with DNMT3A, 1 with TET2, 1with KRAS and 1 with TP53) with a VAF<1.5%, overall resulting in 10patients (27.8%) having detectable somatic CHIP mutations. The expectedfrequency of CHIP in a matched healthy 40-60 group would beapproximately 2-3%. In addition, 5 patients had germline mutations inCHIP genes (3 with TET2, 1 with PP1MD, 1 with TP53). None of the 4samples collected in non-hospitalized COVID19 infected patients in thisage group demonstrated CHIP with a VAF>1.5%.

Age Group >65

This cohort includes 43 hospitalized patients with severe disease, 46non-hospitalized infected patients and a historic control of 106individuals. The median age for each group is 69.5, 68.7, and 82respectively. The frequency of CHIP>1.5% VAF was 14%, 16%, 35%respectively.

TABLE 1 NYBC Study >65 40-60 <40 CHIP > 1.5% CHIP > 1.5% CHIP >1 .5%Severe 6/43 (14%) 3/36 (8.3%) 2/9 (22%) Mild 7/45 (16%) 0/4 (0%) N/AHealthy (33%) (2-3%) (0.1%) PopulationGermline Mutations (IL-6R p. Asp358Ala Coding Mutation)

In addition, multiple studies have demonstrated the increased risk ofcardiovascular disease in the presence of CHIP (17, 27). However, therisk of incident cardiovascular events such as myocardial infarction,stroke and death is abrogated in those with a simultaneous geneticdeficiency of IL-6 signaling (IL-6R p Asp358Ala) (24). It ishypothesized that the increased risk of CVD is mediated by IL-6signaling. Likewise, the severity of COVID-19 has been associated with adysfunctional inflammatory response mediated by cytokines, particularlyIL-6. In one example, a COVID-19 infected patient with two large cloneCHIP mutations (JAK2 mutation with 76% VAF and TET2 mutation with 23%VAF) also had an IL-6R coding mutation which may have protected thispatient from developing severe disease. In this study, 5.1% of patientswith CHIP and a simultaneous genetic deficiency in IL-6 signaling (bycarrying IL6R p.Asp358A1a) were non-hospitalized when infected withCOVID-19. On the other hand, only 1.6% of patients with CHIP and thiscoding mutation developed severe COVID-19 disease. Therefore,genetically reduced IL-6 signaling may abrogate the risk of developingsevere disease among carriers of CHIP.

Third Party Studies

Additional studies looking at the association of CHIP with COVID-19 havebeen conducted by other research groups. A first study is identifiedherein as the French study and described in Duployez, et al., (26), anda second study is identified herein as the MSK study and described inBolton et al., (28). Both studies assessed the relationship betweenclonal hematopoiesis and the severity of a COVID-19 infection. The twostudies are summarized below.

French Study (Duployez, 26):

122 patients hospitalized for COVID19. 76% male. 73% ICU.

A high prevalence of Clonal Hematopoiesis (25%, 38%, 56%, and 82% ofpatients aged <60 years, 60-70 years, 70-80 years, and >80 years) wasreported compared to a retrospective cohort of patients explored withinthe same pipelines (10%, 21%, 37%, and 44%). 80% of those with CHIP hadDNMT3A and/or TET2 mutations. After adjustment for age, the prevalenceOR of CH was 3.182 (95% CI: 1.944-5.209, p<0.001) in COVID-19 patients.

TABLE 2 French Study summary Age CHIP Positive Control Group <60 25% 10%60-70 38% 21% 70-80 56% 37% >80 82% 44%

MSK Study (Bolton, 28):

Among 515 individuals with Covid-19 from Memorial Sloan Kettering (MSK)and the Korean Clonal Hematopoiesis (KoCH) consortia, it was found thatCH was associated with severe Covid-19 outcomes (OR=1.9, 95%=1.2-2.9,p=0.01).

The first cohort was composed of patients with solid tumors treated atMemorial Sloan Kettering Cancer Center (MSK) with blood previouslysequenced using MSK-IMPACT, a previously validated targeted gene panelcapturing all commonly mutated CH-associated genes. In the MSK cohort,CH was observed in 51% and 30% of patients with severe versus non-severeCovid-19, respectively (adjusted OR: 1.85, 95% CI 1.10-3.12).

The second cohort included 112 previously healthy individuals withoutcancer who were hospitalized for Covid-19 between January and April 2020at four tertiary hospitals in South Korea (KoCH cohort). The KoCH cohortwas sequenced using a custom targeted NGS panel from Agilent (89 genes)which was designed to include commonly occurring CH genes.

In the KoCH cohort, CH was observed in 25% and 15.9% of patients withsevere versus non-severe Covid-19, respectively (adjusted OR 1.85, 95%CI 0.53-6.43). A comparative analysis of age groups was not performed.

TABLE 3 MSK Study summary Severe Severe Non-Severe Control Non- OddsChip − CHIP + Chip − Severe Chip + Ratio MSK 46 48 223 96 1.85 KoCH 5117 37 7 1.85

Discussion

The frequency of clonal hematopoiesis is rare in healthy individualsunder 40 and uncommon in those under 60. The results from the studydescribed herein demonstrate there is a relatively high frequency ofCHIP in younger patients infected with COVID-19 who have beenhospitalized with severe disease. And when lower VAFs were included inthe analysis, this frequency increased further. CHIP may provide anexplanation why some younger patients infected with COVID-19 have aclinical course that requires hospitalization. This is consistent withprior studies implicating CHIP in dysfunctional inflammatory responseswith elevated cytokines and decreased interferons which is the typicalprofile of patients with severe disease (REF). Therefore, this would bethe presumed mechanism in the younger cohort with severe disease in ourstudy. Furthermore, acute and chronic cardiovascular events are afrequent cause of morbidity and mortality in hospitalized COVID-19patients and CHIP has been highly associated with cardiovasculardiseases such as myocardial infarction and stroke. Not only is CHIP asignificant risk factor for myocardial infarction of all ages, but therisk is highest in younger patients who demonstrates early-onsetmyocardial infarction (27). Mutations in DNMT3A and TET2 were eachindividually associated with coronary heart disease and in fact, thesewere the most frequently mutated in the CHIP patients with severedisease who were assessed in the study described herein.

The data to date does not show a higher frequency of CHIP inindividuals >65 years old with severe disease compared to the controlgroups as was demonstrated in two recent studies (26, 28). It washypothesized that the presence of other comorbidities in the olderpatients in the study contribute to a similar degree as CHIP and thesecomorbidities are less frequent in younger populations.

Regarding the younger cohort, while there is a paucity of literaturestudying younger individuals with CHIP, a few recent studies haveidentified younger patients with severe autoimmune diseases as havingCHIP. For example, CHIP was found with a relatively high frequency inpatients with RA, but particularly in those with clinically advanced andrefractory disease and including some patients in their 30s (BMJ).

The data provided from this study also demonstrates that geneticallyreduced IL-6 signaling which has been previously shown to be protectivefor cardiovascular insults may abrogate the risk of developing severedisease among carriers of CHIP.

REFERENCES

-   1. Tahamtan, A., and A. Ardebili. 2020. Real-time RT-PCR in COVID-19    detection: issues affecting the results. Expert Rev Mol Diagn 20:    1-2.-   2. Lotfi, M., M. R. Hamblin, and N. Rezaei. 2020. COVID-19:    Transmission, prevention, and potential therapeutic opportunities.    Clin Chim Acta 508: 254-266.-   3. Grant, M. C., L. Geoghegan, M. Arbyn, Z. Mohammed, L.    McGuinness, E. L. Clarke, and R. G. Wade. 2020. The prevalence of    symptoms in 24,410 adults infected by the novel coronavirus    (SARS-CoV-2; COVID-19): A systematic review and meta-analysis of 148    studies from 9 countries. Plos One 15: e0234765.-   4. Lauc, G., and D. Sinclair. 2020. Biomarkers of biological age as    predictors of COVID-19 disease severity. Aging Albany Ny 12:    6490-6491.-   5. Xiao, W.-W., J. Xu, L. Shi, Y.-D. Wang, and H.-Y. Yang. 2020. Is    chronic obstructive pulmonary disease an independent predictor for    adverse outcomes in coronavirus disease 2019 patients? Eur Rev Med    Pharmaco 24: 11421-11427.-   6. Ou, M., J. Zhu, P. Ji, H. Li, Z. Zhong, B. Li, J. Pang, J. Zhang,    and X. Zheng. 2020. Risk factors of severe cases with COVID-19: a    meta-analysis. Epidemiol Infect 148: e175.-   7. Zhang, J., X. Dong, Y. Cao, Y. Yuan, Y. Yang, Y. Yan, C. A.    Akdis, and Y. Gao. 2020. Clinical characteristics of 140 patients    infected with SARS-CoV-2 in Wuhan, China. Allergy 75: 1730-1741.-   8. Huang, C., Y. Wang, X. Li, L. Ren, J. Zhao, Y. Hu, L. Zhang, G.    Fan, J. Xu, X. Gu, Z. Cheng, T. Yu, J. Xia, Y. Wei, W. Wu, X.    Xie, W. Yin, H. Li, M. Liu, Y. Xiao, H. Gao, L. Guo, J. Xie, G.    Wang, R. Jiang, Z. Gao, Q. Jin, J. Wang, and B. Cao. 2020. Clinical    features of patients infected with 2019 novel coronavirus in Wuhan,    China. Lancet 395: 497-506.-   9. Zhou, F., T. Yu, R. Du, G. Fan, Y. Liu, Z. Liu, J. Xiang, Y.    Wang, B. Song, X. Gu, L. Guan, Y. Wei, H. Li, X. Wu, J. Xu, S.    Tu, Y. Zhang, H. Chen, and B. Cao. 2020. Clinical course and risk    factors for mortality of adult inpatients with COVID-19 in Wuhan,    China: a retrospective cohort study. Lancet 395: 1054-1062.-   10. Unudurthi, S. D., P. Luthra, R. J. C. Bose, J. McCarthy,    and M. I. Kontaridis. 2020. Cardiac inflammation in COVID-19:    Lessons from heart failure. Life Sci 260: 118482.-   11. Magadum, A., and R. Kishore. 2020. Cardiovascular Manifestations    of COVID-19 Infection. Cells 9: 2508.-   12. Shivarov, V., and M. Ivanova. 2020. Clonal haematopoiesis and    COVID-19: A possible deadly liaison. Int J Immunogenet 47: 329-331.-   13. Jaiswal, S., P. Fontanillas, J. Flannick, A. Manning, P. V.    Grauman, B. G. Mar, R. C. Lindsley, C. H. Mermel, N. Burtt, A.    Chavez, J. M. Higgins, V. Moltchanov, F. C. Kuo, M. J. Kluk, B.    Henderson, L. Kinnunen, H. A. Koistinen, C. Ladenvall, G. Getz, A.    Correa, B. F. Banahan, S. Gabriel, S. Kathiresan, H. M.    Stringham, M. I. McCarthy, M. Boehnke, J. Tuomilehto, C. Haiman, L.    Groop, G. Atzmon, J. G. Wilson, D. Neuberg, D. Altshuler, and B. L.    Ebert. 2014. Age-Related Clonal Hematopoiesis Associated with    Adverse Outcomes. New Engl J Medicine 371: 2488-2498.-   14. Mas-Peiro, S., J. Hoffmann, S. Fichtlscherer, L.    Dorsheimer, M. A. Rieger, S. Dimmeler, M. Vasa-Nicotera, and A. M.    Zeiher. 2019. Clonal haematopoiesis in patients with degenerative    aortic valve stenosis undergoing transcatheter aortic valve    implantation. Eur Heart J 41: 933-939.-   15. Sano, S., Y. Wang, Y. Yura, M. Sano, K. Oshima, Y. Yang, Y.    Katanasaka, K.-D. Min, S. Matsuura, K. Ravid, G. Mohi, and K.    Walsh. 2019. JAK2 V617F-Mediated Clonal Hematopoiesis Accelerates    Pathological Remodeling in Murine Heart Failure. Jacc Basic Transl    Sci 4: 684-697.-   16. Bazeley, P., R. Morales, and W. H. W. Tang. 2020. Evidence of    Clonal Hematopoiesis and Risk of Heart Failure. Curr Hear Fail    Reports 17: 271-276.-   17. Jaiswal, S., and P. Libby. 2020. Clonal haematopoiesis:    connecting ageing and inflammation in cardiovascular disease. Nat    Rev Cardiol 17: 137-144.-   18. Yura, Y., S. Sano, and K. Walsh. 2020. Clonal Hematopoiesis: A    New Step Linking Inflammation to Heart Failure. Jacc Basic Transl    Sci 5: 196-207.-   19. Abplanalp, W. T., S. Cremer, D. John, J. Hoffmann, B.    Schuhmacher, M. Merten, M. A. Rieger, M. Vasa-Nicotera, A. M.    Zeiher, and S. Dimmeler. 2021. Clonal Hematopoiesis-Driver DNMT3A    Mutations Alter Immune Cells in Heart Failure. Circ Res 128:    216-228.-   20. Fuster, J. J., S. MacLauchlan, M. A. Zuriaga, M. N.    Polackal, A. C. Ostriker, R. Chakraborty, C.-L. Wu, S. Sano, S.    Muralidharan, C. Rius, J. Vuong, S. Jacob, V. Muralidhar, A. A. B.    Robertson, M. A. Cooper, V. Andrés, K. K. Hirschi, K. A. Martin,    and K. Walsh. 2017. Clonal hematopoiesis associated with TET2    deficiency accelerates atherosclerosis development in mice. Science    355: 842-847.-   21. Sano, S., K. Oshima, Y. Wang, S. MacLauchlan, Y. Katanasaka, M.    Sano, M. A. Zuriaga, M. Yoshiyama, D. Goukassian, M. A.    Cooper, J. J. Fuster, and K. Walsh. 2018. Tet2-Mediated Clonal    Hematopoiesis Accelerates Heart Failure Through a Mechanism    Involving the IL-1β/NLRP3 Inflammasome. J Am Coll Cardiol 71:    875-886.-   22. Wang, Y., S. Sano, Y. Yura, Z. Ke, M. Sano, K. Oshima, H.    Ogawa, K. Horitani, K.-D. Min, E. Miura-Yura, A. Kour, M. A.    Evans, M. A. Zuriaga, K. K. Hirschi, J. J. Fuster, E. M. Pietras,    and K. Walsh. 2020. Tet2-mediated clonal hematopoiesis in    non-conditioned mice accelerates age-associated cardiac dysfunction.    Jci Insight 2020 Mar. 26; 5(6).-   23. Rauch, P. J., A. J. Silver, J. Gopakumar, M. McConkey, E.    Sinha, M. Fefer, E. Shvartz, G. Sukhova, P. Libby, B. L. Ebert,    and S. Jaiswal. 2018. Loss-of-Function Mutations in Dnmt3a and Tet2    Lead to Accelerated Atherosclerosis and Convergent Macrophage    Phenotypes in Mice. Blood 132: 745-745.-   24. Bick, A. G., J. P. Pirruccello, G. K. Griffin, N. Gupta, S.    Gabriel, D. Saleheen, P. Libby, S. Kathiresan, and P.    Natarajan. 2019. Genetic IL-6 Signaling Deficiency Attenuates    Cardiovascular Risk in Clonal Hematopoiesis. Circulation 141:    124-131.-   25. Cook, E. K., M. Luo, and M. J. Rauh. 2020. Clonal hematopoiesis    and inflammation: partners in leukemogenesis and comorbidity. Exp    Hematol 83: 85-94.-   26. Duployez, N., J. Demonchy, C. Berthon, J. Goutay, M. Caplan,    A.-S. Moreau, A. Bignon, A. Marceau-Renaut, D. Garrigue, I.    Raczkiewicz, S. Geffroy, M. Bucci, K. Alidjinou, J. Demaret, M.    Labalette, T. Brousseau, A. Dupont, A. Rauch, J. Poissy, S.    Susen, C. Preudhomme, and B. Quesnel. 2020. Clinico-Biological    Features and Clonal Hematopoiesis in Patients with Severe COVID-19.    Cancers (2020) 12(7): 1992.-   27. S. Jaiswal, P. Natarajan, A. J. Silver, C. J. Gibson, A. G.    Bick, E. Shvartz, M. McConkey, N. Gupta, S. Gabriel, D.    Ardissino, U. Baber, R. Mehran, V. Fuster, J. Danesh, P.    Frossard, D. Saleheen, O. Melander, G. K. Sukhova, D. Neuberg, P.    Libby, S. Kathiresan, and B. L. Ebert. Clonal Hematopoiesis and Risk    of Atherosclerotic Cardiovascular Disease N Engl J Med 2017;    377:111-121.-   28. Bolton, K., Youngil Koh, Michael B. Foote, Hogune Im, et al.    Clonal hematopoiesis is associated with risk of severe Covid-19.    medRxiv 2020. 11.25.20233163.-   29. Savola, P., Lundgren, S., Keränen, M. A. I. et al. Clonal    hematopoiesis in patients with rheumatoid arthritis. Blood Cancer    Journal 8, 69 (2018).-   30. Tariq F, Alobaidi B, Xavier M on behalf of Human Dendritic Cell    Lab, Newcastle University, United Kingdom, et al HU0026 CLONAL    HAEMATOPOIESIS ASSOCIATED SOMATIC MUTATIONS IN RHEUMATOID ARTHRITIS    Annals of the Rheumatic Diseases 2020; 79:226.-   31. Tyrrell, D. J., Goldstein, D. R. Ageing and atherosclerosis:    vascular intrinsic and extrinsic factors and potential role of IL-6.    Nat Rev Cardiol 18, 58-68 (2021).-   32. Pedersen K M, Çcolak Y, Ellervik C, et al. Loss-of-function    polymorphism in IL6R reduces risk of JAK2V617F somatic mutation and    myeloproliferative neoplasm: A Mendelian randomization study.    Eclinicalmedicine. 2020 April; 21:100280.

1. A method of treating a subject at risk of developing a severecomplication from Coronavirus disease 2019 (COVID-19) infection, themethod comprising determining whether the subject has clonalhematopoiesis of indeterminate potential (CHIP), wherein the step ofdetermining whether the subject has CHIP comprises sequencing at leastpart of the genome of one or more cells in a blood sample of the subjectto identify a mutation in one or more genes selected from the groupconsisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRAS andSF3B1, wherein the presence of said mutation indicates that the subjectis at risk of developing a severe complication from COVID-19 infection;and treating the subject identified as being at risk of developing asevere complication from COVID-19, wherein treating the subjectcomprises administering to the subject a treatment selected from thegroup consisting of an antiviral medication, monoclonal antibodies, andcombinations thereof to reduce the subject's likelihood of contracting asevere complication from COVD-19 infection.
 2. The method of claim 1,wherein treating the subject further comprises administering to thesubject one or more agents or vaccines to reduce the subject'slikelihood of contracting COVD-19 infection.
 3. The method of claim 1,wherein the subject is under the age of
 40. 4. The method of claim 1,wherein the one or more cells in the blood sample are nucleated cells.5. The method of claim 1, wherein the one or more cells in the bloodsample are somatic cells.
 6. The method of claim 1, wherein the mutationis a somatic mutation in DNMT3A and/or TET2.
 7. The method of claim 1,wherein the mutation is in DNMT3A in exons 7 to
 23. 8. The method ofclaim 1, wherein the mutation is a mis-sense mutation in DNMT3A selectedfrom the group consisting of G543C, S714C, F732C, Y735C, R736C, R749C,F751C, W753C, and L889C.
 9. A method of treating a subject at risk ofdeveloping a severe coronavirus infection comprising determining whetherthe subject has clonal hematopoiesis of indeterminate potential (CHIP),wherein the step of determining whether the subject has CHIP comprisessequencing at least part of the genome of one or more cells in a bloodsample of the subject to identify a mutation in one or more genesselected from the group consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2,TP53, SRSF2, KRAS and SF3B1, wherein the presence of said mutationindicates that the subject has CHIP and is at risk of developing asevere coronavirus infection; and treating the subject having CHIP byadministering a treatment selected from the group consisting of anantiviral medication, monoclonal antibodies, and combinations thereof,to reduce the subject's risk of developing a severe coronavirusinfection.
 10. The method of claim 9, wherein the coronavirus isselected from the group consisting of severe acute respiratory syndromecoronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus(MERS-CoV), and severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).
 11. The method of claim 9, wherein the coronaviruscomprises severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)or COVID-19.
 12. The method of claim 11, wherein treating the subjectfurther comprises administering to the subject one or more agents orvaccines to reduce the subject's likelihood of contracting the severeacute respiratory syndrome coronavirus 2 (SARS-CoV-2) or COVID-19infection.
 13. The method of claim 9, wherein the subject is under theage of
 40. 14. The method of claim 9, wherein the one or more cells inthe blood sample are nucleated cells.
 15. The method of claim 9, whereinthe one or more cells in the blood sample are somatic cells.
 16. Themethod of claim 9, wherein the mutation is a somatic mutation in DNMT3Aand/or TET2.
 17. The method of claim 9, wherein the mutation is inDNMT3A in exons 7 to
 23. 18. The method of claim 9, wherein the mutationis a mis-sense mutation in DNMT3A selected from the group consisting ofG543C, S714C, F732C, Y735C, R736C, R749C, F751C, W753C, and L889C. 19.(canceled)
 20. (canceled)
 21. A method of selecting hematopoietic cellsfor transplantation, the method comprising the steps of: (a) obtaining asample comprising hematopoietic cells from a human subject; (b)sequencing at least part of the genome of one or more cells in thesample to identify a mutation in one or more genes selected from thegroup consisting of DNMT3A, TET2, ASXL1, PPM1D, JAK2, TP53, SRSF2, KRASand SF3B1; and (c) collecting the remaining hematopoietic cells in thesample for transplantation.